Cyclin-dependent kinases (Cdks) are serine-threonine protein kinases that play critical roles in regulating the transitions between different phases of the cell-cycle, such as the progression from a quiescent stage in G1 (the gap between mitosis and the onset of DNA replication for a new round of cell division) to S (the period of active DNA synthesis), or the progression from G2 to M phase, in which active mitosis and cell-division occurs. (See, e.g., the articles compiled in Science, 274:1643-1677 (1996); and Ann. Rev. Cell Dev. Biol., 13:261-291 (1997)). CDK complexes are formed through association of a regulatory cyclin subunit (e.g., cyclin A, B1, B2, D1, D2, D3 and E) and a catalytic kinase subunit (e.g., CDK1, CDK2, CDK4, CDK5 and CDK6). As the name implies, the CDKs display an absolute dependence on the cyclin subunit in order to phosphorylate their target substrates, and different kinase/cyclin pairs function to regulate progression through specific phases of the cell-cycle.
As seen above, these protein kinases are a class of proteins (enzymes) that regulate a variety of cellular functions. This is accomplished by the phosphorylation of specific amino acids on protein substrates resulting in conformational alteration of the substrate protein. The conformational change modulates the activity of the substrate or its ability to interact with other binding partners. The enzyme activity of the protein kinase refers to the rate at which the kinase adds phosphate groups to a substrate. It can be measured, for example, by determining the amount of a substrate that is converted to a product as a function of time. Phosphorylation of a substrate occurs at the active-site of a protein kinase.
Because CDKs such as CDK1 serve as general activators of cell division, inhibitors of CDK1 can be used as antiproliferative agents. These inhibitors can be used for developing therapeutic intervention in suppressing deregulated cell cycle progression.
It is desirable to provide small molecule inhibitors of Cdk1 that are selective against other Cdks. That is, the small molecule is significantly more inhibitory of Cdk1 activity than Cdk2 and/or Cdk4 activity. Preferably, the compounds of the invention are at least two times, most preferably ten times, more inhibitory of Cdk1 activity than Cdk2 activity and at least five hundred times, preferably one thousand times, more inhibitory of Cdk1 activity than Cdk4 activity. Selectivity is believed to be a desirable parameter because of the potential concomitant toxicity and other undesirable complications that may follow from inhibiting multiple targets. Thus, for purposes of this invention, the inhibition of Cdk2 and Cdk4 are monitored to determine the selectivity of the inhibition of Cdk1. A compound that exhibits selectivity against Cdk2 and Cdk4 is expected to have a better safety profile than a compound that is not selective between Cdk1, Cdk2, and Cdk4.